Cell Therapy For Parkinson's Disease Research Articles

Clinical considerations in Parkinson's disease cell therapy.

Cell replacement therapy is an area of increasing interest for treating Parkinson's disease (PD). However, to become a clinically practical option for PD patients, it must first overcome significant barriers, including establishment of safe and standardized surgical procedures, determination of appropriate perioperative medication regimens, demonstration of long-term graft survival and incorporation, and standardized, clinically meaningful follow-up measures. In this review, we will describe the current status of cell therapy for PD with special attention to these critical requirements, to define guideposts on the road to bring the benefit of this therapy to the Parkinson's clinic.

A new area of stem cell therapy for Parkinson disease

No treatment currently can be used in order to slow or even stop the progression of Parkinson's disease. Nowadays, researchers are already using stem cells to grow dopamine-producing nerve cells in the lab so that they can study the disease, especially in those cases where there is a known genetic cause for the condition. The development of the advanced cellular therapies and using induced pluripotent stem cells is making it possible to combat the progression of the disease without the resulting motor complications. It has been shown that the transplantation of many cell sources leads to reduce Parkinson’s disease symptoms in animal models.

Mitomycin-C treatment during differentiation of induced pluripotent stem cell-derived dopamine neurons reduces proliferation without compromising survival or function in vivo.

Nongenetic methodologies to reduce undesirable proliferation would be valuable when generating dopamine neurons from stem cells for transplantation in Parkinson's disease (PD). To this end, we modified an established method for controlled differentiation of human induced pluripotent stem cells (iPSCs) into midbrain dopamine neurons using two distinct methods: omission of FGF8 or the in‐process use of the DNA cross‐linker mitomycin‐C (MMC). We transplanted the cells to athymic rats with unilateral 6‐hydroxydopamine lesions and monitored long‐term survival and function of the grafts. Transplants of cells manufactured using MMC had low proliferation while still permitting robust survival and function comparable to that seen with transplanted dopamine neurons derived using genetic drug selection. Conversely, cells manufactured without FGF8 survived transplantation but exhibited poor in vivo function. Our results suggest that MMC can be used to reduce the number of proliferative cells in stem cell‐derived postmitotic neuron preparations for use in PD cell therapy.

PCL/gelatin scaffolds and beta-boswellic acid synergistically increase the efficiency of CGR8 stem cells differentiation into dopaminergic neuron: A new paradigm of Parkinson's disease cell therapy.

Parkinson's disease is a progressive degenerative disorder in the central nervous system, which is distinguished by the death of dopamine-producing nerve cells. Levodopa, a dopamine precursor drug, is the current standard of care of symptomatic treatment for Parkinson's disease. However, the long-term use of the drug is associated with the development of motor fluctuations and dyskinesias. Cellular therapies aim to deploy fetal dopaminergic neurons as a means to replace the missing dopamine-producing cells. The present study aims to study the impact of beta-boswellic acid (BBA) coupled with poly ε-caprolactone (PCL)/gelatin scaffolds on the dopaminergic differentiation course of CGR8 embryonic stem cells (ESCs). For this purpose, CGR8 ESCs were cultured on PCL/gelatin scaffolds and a five-step protocol was employed to be promoted the neural differentiation of CGR8 ESCs. Gene expression analysis by real-time qPCR demonstrated that PCL/gelatin scaffolds along with BBA treatment impose synergistic effects on the derivation of dopaminergic-like cells from CGR8 ESCs. Reverse-phase high-performance liquid chromatography confirmed the functionality of the derived neurons by demonstrating the efficient secretion of dopamine in response to stimuli. Our results suggested that the generation of functional dopaminergic-like cells from CGR8 ESCs was increased and supported by PCL/gelatin scaffolds and BBA treatment can heighten the efficiency. The result of this study may open insight into Parkinson's disease cell therapy and provide future directions for tissue engineering aimed at the treatment of Parkinson's disease.

C-Stem, a new technology fro scale up of cell therapy production

Background & Aim Most stem cells, including Human pluripotent stem cells (hPSCs) are adherent cells. The historical standard of culture is in 2D at the bottom of Petri dishes. This technology allowed for the first trials in cell therapy but is limited by: i) reproducibility issues, ii) scalability, iii) high cost due to labor intensive manipulation of the cells, and iv) genomic instability. For cell therapy applications, cell quality reflects the culture process and large quantities of cells are required for allogenic strategies. 2D automation enables reproducible clinical batch production but commercial batch sizes will require further scale up. Suspension culture in bioreactors is a straightforward choice for scale up with two main techniques: i) microcarriers, where the cells grow attached to their surface or ii) spheroids, where cells grow in 3D aggregates. In both culture conditions the cells’ exposure to shear stress in the bioreactors limits further scale up. Methods, Results & Conclusion The C-Stem™ technology addresses this challenge by encapsulating cells within hollow alginate hydrogel capsules, 250 µm in diameter with 30 µm thick walls. Capsules both protect cells form shear stress and let small molecules (smaller than 150 kDa), gas, nutrients and waste in and out. Inside the capsule, hPSCs are seeded in extracellular matrix to mimic in vivo conditions. The capsules are produced at high throughput (5000 per second) with proprietary microfluidics. The C-Stem™ technology allows for 100x fold expansion per week of hPSCs and their differentiation in traditional bioreactors. At the PSC stage, C-Stem™ significantly improves cell viability as compared to standard 2D PSC culture. C-Stem™ promotes pluripotency, yields biomimetic hPSC “epiblastoids” and maintains state of the art genomic integrity. Addressing current manufacturing issues, the process is fully integrated from seeding to harvest, scalable, single use, closed and aligned with cGMP requirements. Most 2D and 3D protocols can be transposed in C-Stem™. Preclinical results will be presented for Parkinson's disease cell therapy. Fully mature dopaminergic neuronal microtissues are differentiated from hPSCs within capsules. They are then engrafted in Parkinson's disease rat model striata. This results in behavioral correction of rats within 6 weeks of surgery, thereby reducing by a factor 2-3 the recovery time as compared to standard progenitor engraftment.

Selection of Cells for Parkinson's Disease Cell-Therapy

Availability of DOPA <em>in situ</em> and thereafter Dopamine (DA) is the main therapeutic approach to treat the Parkinson's Disease (PD). Human neural stem cells (hNSCs), a good source of DA synthesis and release, have long been demonstrated to be a promising candidate for treating PD. However limited cell sourcing and low-growth rate are major concern of its applicability.

Cell therapy for Parkinson's disease is coming of age: current challenges and future prospects with a focus on immunomodulation.

Parkinson's disease (PD) is a neurodegenerative disease that affects more than 1% of people over the age of 60. The principal feature of this disease is the progressive loss of dopaminergic neurons (DAn) within the nigrostriatal system, causing the motor symptoms observed in these patients. At present, there is no therapeutic approach with a cytoprotective effect that can prevent DAn cell death or disease progression. Cell replacement therapy began 30 years ago with the objective to compensate for the loss of DAn by transplantation of dopamine-producing cells. The results from these trials have provided proof of concept of safety and efficacy of cell replacement. However, a major limiting factor of this strategy has been the poor survival rate of grafted DAn. An important factor that could cause cell death of DA precursors is the host response to the graft. In this review, we discuss the factors that affect the outcome of cell therapy in PD, with focus on the cell types used and the functional effects of the host immune response on graft survival and differentiation. We also discuss the strategies that may increase the efficacy of cell replacement therapy which target the host immune response.

Effects of Rho Kinase Inhibitors on Grafts of Dopaminergic Cell Precursors in a Rat Model of Parkinson's Disease.

In models of Parkinson's disease (PD), Rho kinase (ROCK) inhibitors have antiapoptotic and axon-stabilizing effects on damaged neurons, decrease the neuroinflammatory response, and protect against dopaminergic neuron death and axonal retraction. ROCK inhibitors have also shown protective effects against apoptosis induced by handling and dissociation of several types of stem cells. However, the effect of ROCK inhibitors on dopaminergic cell grafts has not been investigated. In the present study, treatment of dopaminergic cell suspension with ROCK inhibitors yielded significant decreases in the number of surviving dopaminergic neurons, in the density of graft-derived dopaminergic fibers, and in graft vascularization. Dopaminergic neuron death also markedly increased in primary mesencephalic cultures when the cell suspension was treated with ROCK inhibitors before plating, which suggests that decreased angiogenesis is not the only factor leading to cell death in grafts. Interestingly, treatment of the host 6-hydroxydopamine-lesioned rats with ROCK inhibitors induced a slight, nonsignificant increase in the number of surviving neurons, as well as marked increases in the density of graft-derived dopaminergic fibers and the size of the striatal reinnervated area. The study findings discourage treatment of cell suspensions before grafting. However, treatment of the host induces a marked increase in graft-derived striatal reinnervation. Because ROCK inhibitors have also exerted neuroprotective effects in several models of PD, treatment of the host with ROCK inhibitors, currently used against vascular diseases in clinical practice, before and after grafting may be a useful adjuvant to cell therapy in PD. Cell-replacement therapy is one promising therapy for Parkinson's disease (PD). However, many questions must be addressed before widespread application. Rho kinase (ROCK) inhibitors have been used in a variety of applications associated with stem cell research and may be an excellent strategy for improving survival of grafted neurons and graft-derived dopaminergic innervation. The present results discourage the treatment of suspensions of dopaminergic precursors with ROCK inhibitors in the pregrafting period. However, treatment of the host (patients with PD) with ROCK inhibitors, currently used against vascular diseases, may be a useful adjuvant to cell therapy in PD.

Directed Differentiation of Dopamine-Secreting Cells from Nurr1/GPX1 Expressing Murine Embryonic Stem Cells Cultured on Matrigel-Coated PCL Scaffolds.

Parkinson's disease (PD) is a progressive neurological disorder characterized by a large number of motor and non-motor features and is known as the second most common neurodegenerative disorder after Alzheimer's disease. The hallmark pathology of PD is the damage and death of dopamine-producing neurons in the substantia-nigra of midbrain. Intrastriatal transplants of fetal mesencephalon derived DAergic neurons have provided proof-of-principle for the cell replacement strategy and have demonstrated reinnervation of the denervated striatum. However, ethical, technical, and practical limitations of deploying fetal DAergic neurons as the source for cell therapy in PD have ceased the spread of this procedure into clinical practice. Embryonic stem (ES) cells have emerged as a therapeutic alternative that can proliferate extensively and generate dopamine-producing neurons. To this extent and to surmount the obstacles related to embryonic neural cells, many investigations have focused on using pluripotent stem cells for the derivation of DAergic neurons. In the present study, a mouse embryonic stem (mES) R1 cell line was generated which could stably co-express Nurr1 (an essential transcription factor in DAergic neuron development) and GPX-1 (a neuroprotective enzyme against oxidative stress). The Nurr1/GPX-1-expressing ES cells (Nurr1/GPX-1-ES) were differentiated into DAergic-like cells via a three-dimensional culture environment consisting of Poly-ε-Caprolactone (PCL) nanofibrous scaffolds embedded by Matrigel (Mtg) in the presence of specific signaling molecules. DAergic neuron-specific genes were highly expressed in ES-derived DAergic neurons cultured and differentiated on PCL/Mtg scaffolds. Reverse-phase HPLC confirmed that the Nurr1/GPX-1-ES-cells differentiated on PCL/Mtg electrospun scaffolds could efficiently and exclusively secrete dopamine in response to stimulus. In conclusion, our results demonstrated that PCL/Matrigel nanofibrous scaffolds could efficiently support and promote the generation of functional DAergic-like cells from Nurr1/GPX-1-ES cells. The results of this study may have an impact on future tissue engineering for cell therapy of PD.

Terapia celular en la enfermedad de Parkinson y los factores que influyen en su éxito

Objective: To determine the factors that influence improvement of patients with Parkinson's disease treated with autologous adult stem cells from bone marrow by intra-arterial infusion. Material and Methods: An observational, analytical and retrospective study. We evaluated 58 patients with Parkinson's disease (PD) who received cell therapy at the Brazzini Institute of Interventional Radiology, between July 2006 and March 2010. The improvement was determined by comparing the UPDRS values before the procedure and the best values of patient monitoring. SPSS version 15.0 was used for analysis of relationship between improvement and intrinsic factors of the patient and his disease. P &lt;0.05 to determine statistical significance. Results: 62% of patients achieved a good improvement in UPDRS values (more than 50% improvement). No significant differences (p&gt; 0.05) in the results of improvement were found by gender. Significant differences (p &lt;0.05) in the improvement according to age groups. Patients with more improvement were in the group of 50-59 years and those with little improvement were in the group of 70-79 years. Intrinsic factors such as severe dyskinesia, hypertension, diabetes mellitus, cancer in remission, family history of PD and month of follow-up did not influence the result of improvement (p&gt; 0.05). Conclusion: Factors influencing the improvement in patients treated with cell therapy for PD are patient's age at disease onset and age at treatment. We recommend using these factors to choose patients who receive this therapy.

Identification of Neurexophilin 3 as a Novel Supportive Factor for Survival of Induced Pluripotent Stem Cell-Derived Dopaminergic Progenitors.

Successful cell transplantation for Parkinson's disease (PD) depends on both an optimal host brain environment and ideal donor cells. We report that a secreted peptide, neurexophilin 3 (NXPH3), supports the survival of mouse induced pluripotent stem cell-derived (iPSC-derived) dopaminergic (DA) neurons in vitro and in vivo. We compared the gene expression profiles in the mouse striatum from two different environments: a supportive environment, which we defined as 1 week after acute administration of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), and a nonsupportive environment, defined as 8 weeks after chronic administration of MPTP. NXPH3 expression was higher in the former condition and lower in the latter compared with untreated controls. When we injected mouse iPSC-derived neural cells along with NXPH3 into the mouse striatum, the ratio of tyrosine hydroxylase-positive DA neurons per graft volume was higher at 8 weeks compared with cell injections that excluded NXPH3. In addition, quantitative polymerase chain reaction analyses of the postmortem putamen revealed that the expression level of NXPH3 was lower in PD patients compared with normal controls. These findings will contribute to optimizing the host brain environment and patient recruitment in cell therapy for PD.

Direct pre-differentiation of rat mesenchymal stem cells into dopaminergic cells

The use of stem cells has been proposed as an alternative treatment for certain neurodegenerative disorders. It has also been suggested that in the pre-differentiated state, stem cells might provide a better therapeutic option than cells that are undifferentiated or fully differentiated. The purpose of this study was to develop a protocol aimed at reducing the incubation time required to induce the conversion of rat mesenchymal stem cells into immature dopaminergic neurons. Stem cells obtained from rat bone marrow were incubated in a control or induction media for 2 - 24 h. Cells incubated for 24 h in induction medium demonstrated an increase on the levels of the neuronal protein markers nestin, glial fibrillary acid protein, and β-tubulin III, as well as increases in the expression of Pax3, EN1, Thy1.1, and GEF10 genes. This manuscript presents evidence that adult mesenchymal cells are capable to respond, in a short time period, to a neuroinduction medium, and give raise to predifferentiated neuron like cells representing an alternative for Parkinson disease cell therapy transplantation.

Stem cells and regenerative therapies for Parkinson's disease.

Currently the mainstay of Parkinson's disease (PD) therapy is the pharmacological replacement of the loss of the dopaminergic nigrostriatal pathway using drugs such as dopamine agonists and levodopa. Whilst these drugs effectively ameliorate some of the motor features of PD, they do not improve many of the nonmotor features that arise secondary to pathology outside of this system, nor do they slow the progressive neurodegeneration that is a characteristic of the disease. Regenerative therapies for PD seek to fill this therapeutic gap, with cell transplantation being the most explored approach to date. A number of different cell sources have been used in this therapeutic approach, but to date, the most successful has been the use of fetal ventral mesencephalic (VM) tissue that contains within it the developing nigral dopaminergic cells. Cell transplantation for PD was pioneered in the 1980-1990s, with several successful open-label trials of fetal VM transplantation in patients with relatively advanced PD. Whilst these findings were not replicated in two subsequent double-blind sham-surgery controlled trials, there were reasons to explain this outside of the one drawn at the time that these therapies are ineffective. Indeed all these studies have provided evidence that following the transplantation of fetal VM tissue, dopaminergic cells can survive long term, produce dopamine, and bring about clinical improvements in younger patients over many years. The use of fetal tissue, irrespective of its true efficacy, will never become a widely available therapy for PD for a host of practical and ethical reasons, and thus much work has been put in recently to exploring the utility of stem cells as a source of nigral dopaminergic neurons. In this respect, the advent of embryonic stem cell and induced pluripotent cells has heralded a new era in cell therapy for PD, and several groups have now demonstrated that these cells can form dopaminergic neurons which improve functional deficits in animal models of PD. Whilst encouraging, problems with respect to the immunogenicity and tumorigenicity of these cells means that they will need to be used in the clinic cautiously. Other regenerative therapies in PD have been tried over the years and include the use of trophic factors. This has primarily involved glial cell line-derived neurotrophic factor (GDNF) and again has produced mixed clinical effects, and in order to try and resolve this, a new trial of intraputamenal GDNF is now being planned. In addition, a new trial for platelet derived growth factor as a treatment for PD has just completed recruitment, and PYM50028 (Cogane) an oral agent shown in animal models to reduce the effects of MPTP (1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine) lesioning by the induction of growth factors is currently under investigation in a multicentre Phase II trial. Overall, there are a number of promising new regenerative therapies being developed and tested in PD, although the true long-term efficacy of any of these in large numbers of patients is still not known.

Impact of dopamine versus serotonin cell transplantation for the development of graft-induced dyskinesia in a rat Parkinson model

Graft-induced dyskinesia (GID), covering a range of dystonic and choreiform involuntary movements, has been observed in some patients with Parkinson's disease (PD) after intracerebral cell transplantation. These dyskinesias have been severe in a number of patients and represent one of the main obstacles for further development of the cell therapy in PD. Serotonin neurons, included into the dopaminergic cell suspension due to the nature of the dissection process, have been suggested as a key factor for the development of GID, since the administration of the serotonin (5-HT)1A-receptor agonist buspirone reduced dyskinesia in transplanted PD patients. In the present study, we characterized GID in the rat PD model after transplantation of dopaminergic grafts containing different amounts of serotonin neurons. The severity of GID was significantly correlated with the amount of grafted dopamine and serotonin neurons, but the r-values were low. However, neither the innervation density of dopamine and serotonin fibers in the grafted striatum nor the dopamine-to-serotonin cell ratio correlated significantly with the severity of GID. The results extend prior knowledge of the role of dopaminergic grafts in the development of GID and show that, in the animal model, serotonin neurons within the graft suspension might be involved, but given sufficient dopamine cells, their impact on GID may be minor.

Cells therapy for Parkinson’s disease—so close and so far away

One of the strategies of treating Parkinson's disease (PD) is the replacement of lost neurons in the substantia nigra with healthy dapamingergic cells. Potential sources for cells range from autologous grafts of dopamine secreting cells, fetal ventral mesencephalon tissue, to various stem cell types. Over the past quarter century, many experimental replacement therapies have been tried on PD animal models as well as human patients, yet none resulted in satisfactory outcomes that warrant wide applications. Recent progress in stem cell biology has shown that nuclear transfer embryonic stem cells (ntES) or induced pluripotent stem cells (iPS) derived cells can be used to successfully treat rodent PD models, thus solving the problem of immunorejection and paving the way for future autologous transplantations for treating PD. Meanwhile, however, post mortem analysis of patients who received fetal brain cell transplantation revealed that implanted cells are prone to degeneration just like endogenous neurons in the same pathological area, indicating long-term efficacy of cell therapy of PD needs to overcome the degenerating environment in the brain. A better understanding of neurodegeneration in the midbrain appeared to be a necessary step in developing new cell therapies in Parkinson's disease. It is likely that future cell replacement will focus on not only ameliorating symptoms of the disease but also trying to slow the progression of the disease by either neuroprotection or restoring the micro-environment in the midbrain.

Chemically defined sequential culture media for TH+ cell derivation from human embryonic stem cells

During the past few years several differentiation protocols to derive midbrain dopamine (DA) neurons from human embryonic stem (hES) cells have been developed, but the production of sufficient amounts of the 'right' therapeutic DA cells has not yet been accomplished. The aim of this study was to efficiently generate tyrosine hydroxylase (TH)-positive cells in vitro from our hES cells using a chemically defined culture system. At the end of differentiation, the vast majority of cells (>90%) were positive for both TH and beta-tubulin isotype III (TuJ1). Other markers of dopaminergic cells, like dopamine transporter (DAT) and Nurr1 were also detected by immunofluorescence or RT-PCR. The functions of these cells were confirmed by measurements of DA release in vitro and by transplantation of derived cells into Parkinson's disease (PD) rats in vivo. We found these cells were able to release DA when depolarized by high K(+). Moreover, 4 weeks after transplantation, the hES-derived cells could survive and reduce the apomorphine-induced rotation behaviour of the rats. In conclusion, the experimental system presented here provided a reliable protocol to produce a large number of hES-derived TH(+) cells which may be used in cell therapy for PD in future.

Recent advances in cell-based therapy for Parkinson disease

In this review, the authors discuss recent advances in the field of cell therapy for Parkinson disease (PD). They compare and contrast recent clinical trials using fetal dopaminergic neurons. They attribute differences in cell preparation techniques, cell type specification, and immunosuppression as reasons for variable outcome and for some of the side effects observed in these clinical trials. To address ethical, practical, and technical issues related to the use of fetal cell sources, alternative sources of therapeutic dopaminergic neurons are being developed. The authors describe the progress in enrichment and purification strategies of stem cell-derived dopaminergic midbrain neurons. They conclude that recent advances in cell therapy for PD will create a viable long-term treatment option for synaptic repair for this debilitating disease.

Human Retinal Pigment Epithelial Cell Implants Ameliorate Motor Deficits in Two Rat Models of Parkinson Disease

Intrastriatal transplantation of gelatin microcarrier-attached human retinal pigment epithelial cells (hRPE-GM) may represent an alternative source for cell therapy in Parkinson disease (PD). The use of human retinal pigment epithelial (hRPE) cells in PD relies on the capacity of these cells to produce l-dopa as an intermediate product in the eumelanin synthesis pathway. We investigated the behavioral effects of hRPE-GM implants on forelimb use asymmetries and hindlimb motor deficits in unilateral and bilateral 6-hydroxydopamine (6-OHDA) rat models of PD. We report that intrastriatal unilateral implantation of hRPE-GM in rats with 6-OHDA nigrostriatal lesions produce an amelioration of the contralateral forelimb disuse and the contralateral hindlimb deficits. These results further support the possibility that implantation of cultured hRPE cells may be a promising therapeutic option for patients with PD.

Intracerebral Transplantation of Genetically Engineered Cells for Parkinson's Disease: Toward Clinical Application

Over the last decade, molecular biology has progressively developed, leading to new technology with subsequent clinical application for various cerebral diseases including Parkinson's disease (PD), one of the most investigated neurodegenerative disorders. The therapy for PD is mainly composed of medication, including drug replacement therapy, surgical treatment, and cell transplantation. Cell therapy for PD has been explored by using fetal nigral cells as an allo- or xenograft, autologous sympathetic ganglion, adrenal medulla, and carotid body in clinical settings. In addition, neurotrophic factors, including glial cell line-derived neurotrophic factor (GDNF), have a strong potency to rescue degenerating dopaminergic cells. Protein and/or gene therapy also might be a therapeutic option for PD. In this review, genetically engineered cell transplantation for animal models of PD, including catecholamine/neurotrophic factor-secreting cell transplantation with or without encapsulation, as performed in our laboratories, and their potential future as clinical applications are described with recent clinical studies in this field.

Enhanced Yield of Neuroepithelial Precursors and Midbrain-Like Dopaminergic Neurons from Human Embryonic Stem Cells Using the Bone Morphogenic Protein Antagonist Noggin

It is currently not known whether dopamine (DA) neurons derived from human embryonic stem cells (hESCs) can survive in vivo and alleviate symptoms in models of Parkinson disease (PD). Here, we report the use of Noggin (a bone morphogenic protein antagonist) to induce neuroectodermal cell development and increase the yield of DA neurons from hESCs. A combination of stromal-derived inducing activity and Noggin markedly enhanced the generation of neuroepithelial progenitors that could give rise to DA neurons. In addition, Noggin diminished the occurrence of a fibroblast-like Nestin-positive precursor population that differentiated into myocytes. After transplantation of differentiated hESCs to a rodent model of PD, some grafts contained human midbrain-like DA neurons. This protocol demonstrates hESC derivation and survival of human DA neurons appropriate for cell therapy in PD.